The lymphatic system plays the major role in tissue fluid homeostasis by draining the interstitial fluid back to the circulation. Lymphedema, caused by lymphatic malformation or obstruction, is often associated with radiation and surgery; however effective treatments that address the underlying molecular pathology are not available to date. We have recently reported that 9-cis retinoic acid (RA) can activate cell proliferation, migration and tube formation of lymphatic endothelial cells (LECs), stimulate lymphangiogenesis in vivo, and ameliorate secondary lymphedema by promoting lymphatic regeneration in a mouse model. These pro-lymphangiogenic features of 9-cisRA, however, are quite unexpected, because RAs have been known for their anti-proliferative effects on many cell types, including blood vascular endothelial cells (BECs); where RAs have been shown to suppress BEC proliferation, and RA-deficient mouse embryos display hyper-proliferation of BECs. In this proposal, therefore, we aim to address two main questions (1) what is the molecular mechanism underlying RA-induced lymphangiogenesis, and (2) how can RAs selectively induce lymphangiogenesis, while concurrently suppressing angiogenesis. Our preliminary studies revealed that RAs may regulate Notch pathway to promote lymphatic sprouting, suggesting novel crosstalk between the two important morphogenic signals, and also that Prox1, the master regulator of lymphatic differentiation and development, can physically and functionally interact with a RA-binding nuclear receptor RXR in a RA- controlled manner. Furthermore, LECs predominantly express FABP4 as a cytoplasmic RA-carrier, and PPAR? as a dimerization partner of RXR, which is known to promote cell proliferation in response to RAs, whereas BECs selectively express CRABP-II and RAR?, a molecular pairing that induces cell growth arrest in response to RAs. Together, we propose working hypotheses addressing our two main questions that (1) RAs stimulate lymphatic sprouting by modulating Notch pathway genes through regulation of the interactions of Prox1 and RXR in LECs and (2) the predominant expression of FABP4 and PPAR? in LECs converts RA from an anti-proliferative signal to a pro-growth cue in LECs. Here, we aim to validate these working hypotheses by studying the role of RAs in promoting lymphangiogenesis through RXR? and PPAR? (Aim1), mechanism underlying the opposing effects of RAs on angiogenesis vs. lymphangiogenesis (Aim 2), and RA-controlled physical and functional interactions between Prox1 and RXR? (Aim 3). Together, our studies will not only provide important information on how Prox1 functions as the master regulator of lymphatic development by functioning as a nuclear receptor coregulator, but also define the molecular mechanism underlying RA-mediated selective promotion of lymphangiogenesis. In the long run, our study will help lay an essential experimental foundation to repurpose RAs as potential therapeutic agents for lymphatic circulation insufficiency.